Cardiovascular disease is the leading cause of mortality in the United States, with associated healthcare costs exceeding $200 billion per year. Cardiovascular diseases, including heart disease, commonly culminate in heart failure, in which the heart is incapable of pumping a sufficient supply of blood to the peripheral tissues and organs. Indeed, heart failure has a 50% 5 year survival rate and is the most common hospital discharge diagnosis. Current standards of care include drugs that target pathways involved in the progression of heart disease, such as neuroendocrine stimulation and hypertension, resulting in delayed progression of heart failure and mortality but not affecting the underlying cause of the disease or substantially altering ultimate disease outcomes. Therefore, a better understanding of the molecular pathways directly involved in the pathogenesis of heart disease or that mediate cardioprotection from disease-inducing stimuli (i.e. myocardial infarction) will ai in the development of novel, efficacious therapeutic strategies to treat heart disease. To that end, the sponsor's laboratory has been investigating the role of thrombospondins, a family of calcium-binding glycoproteins with fundamental roles in wound healing and tissue repair, in protection from heart disease. Thrombospondins are induced in cardiac disease states and overexpression of thrombospondin-4 protects the heart from pressure overload hypertrophy (PO) and myocardial infarction injury (MI), while mice lacking thrombospondin-1, 2, or 4 exhibit greatly increased mortality after cardiac stress. The sponsor's laboratory has recently identified a cardioprotective adaptive endoplasmic reticulum (ER) stress pathway that is activated by thbrombospondin-4 (Thbs4) in cardiomyocytes. Overexpression of Thbs4 also results in vesicular expansion and enhanced secretion in the heart and skeletal muscle, suggesting a tissue autonomous role for thrombospondins in regulating flux through the secretory pathway. Moreover, protein interaction studies reveal that Thbs-4 interacts with a number of proteins involved in trafficking of substrates within the cell or to the extracelluar matrix (ECM) or cell membrane. These findings suggest that Thbs4 has critical roles in trafficking substrates to their ultimate destinations to maintain cardiomyocyte homeostasis and/or ECM production. Thus, Thbs4 activates a secretory pathway that may be critical in maintaining normal turnover and localization of proteins within cardiomyocytes and adaptive under cardiac stress conditions, such as PO or MI. Therefore, this proposal will examine 1) the domains of Thbs4 that mediate its functions in ER stress sensing, trafficking, or regulation of ECM production/homeostasis and 2) the roles of Thbs4 in trafficking proteins to intracellular, membranous, and extracellular destinations. These studies will elucidate mechanisms of cellular trafficking and regulation of the ER stress response and provide insight into molecular pathways involved in protection from cardiac disease, thereby aiding in the development of novel therapeutic strategies to treat heart disease and other diseases associated with protein quality control and ECM remodeling.